This invention relates generally to the fields of electroporation and electrofusion, and more specifically to electroporation and electrofusion using low level electric fields.
Electroporation involves the opening of the cell membrane, usually to allow genetic or other material to pass into or out of the cell. When genes are employed, this activity is known as genetic transfection.
Electrofusion, or cell-cell joining, involves the fusion of membranes of different cells after their membranes have been opened by electroporation. Electrofusion is used to form new cells (hybridomas) with unique properties, or to genetically reprogram existing cells such as for plant breeding or genetic engineering.
Conventional methods of opening cell membranes for transfection or cell fusion use inactivated viruses or chemicals such as polyethylene glycol (PEG). These techniques have certain disadvantages. Strict controls are needed in the case of viral transfection to prevent unwanted contamination. Also there are problems with unwanted biologic responses. Furthermore the chemical side effects of PEG transfection or fusion can adversely affect cellular viability.
Electric-field induced fusion, or electrofusion, has shown promising results. In electrofusion, different types of cells are placed in close contact by applying non-uniform alternating current electric fields to a solution of those cells. The electric fields cause dielectrophoresis, which in turn causes cells to move to a region of highest field intensity and organize into formations of variable length.
Once close cell-to-cell membrane contact is achieved, fusion occurs by subjecting the cells to one or more pulses of high intensity electric fields. The high intensity fields first cause reversible electric breakdown in the zone of contact between the two cells, and then fusion of the two cells occurs.
This process, however, also has some drawbacks. Conventional electrofusion employs an unnatural chemical environment in the low conductivity solutions. Also, some conventional electrofusion produces random electrofusion because of the use of cell suspensions. In electrofusing cells A & B, the resulting fused cells could be composed of A:A, B:B, A:A:B, A:B:B, etc., as well as the desired result of simply A:B.
It is therefore an object of this invention to provide methods and apparatus for electroporation and electrofusion which yield more predictable results than conventional methods.
Another object of this invention is to provide methods and apparatus for electroporation and electrofusion which minimize damage caused to the cells.